Methods for the extraction, storage, amplification and sequencing of DNA from environmental samples

Advances in the sequencing of DNA extracted from media such as soil and water offer huge opportunities for biodiversity monitoring and assessment, particularly where the collection or identification of whole organisms is impractical. However, there are myriad methods for the extraction, storage, amplification and sequencing of DNA from environmental samples. To help overcome potential biases that may impede the effective comparison of biodiversity data collected by different researchers, we propose a standardised set of procedures for use on different taxa and sample media, largely based on recent trends in their use. Our recommendations describe important steps for sample pre-processing and include the use of (a) Qiagen DNeasy PowerSoil® and PowerMax® kits for extraction of DNA from soil, sediment, faeces and leaf litter; (b) DNeasy PowerSoil® for extraction of DNA from plant tissue; (c) DNeasy Blood and Tissue kits for extraction of DNA from animal tissue; (d) DNeasy Blood and Tissue kits for extraction of DNA from macroorganisms in water and ice; and (e) DNeasy PowerWater® kits for extraction of DNA from microorganisms in water and ice. Based on key parameters, including the specificity and inclusivity of the primers for the target sequence, we recommend the use of the following primer pairs to amplify DNA for analysis by Illumina MiSeq DNA sequencing: (a) 515f and 806RB to target bacterial 16S rRNA genes (including regions V3 and V4); (b) #3 and #5RC to target eukaryote 18S rRNA genes (including regions V7 and V8); (c) #3 and #5RC are also recommended for the routine analysis of protist community DNA; (d) ITS6F and ITS7R to target the chromistan ITS1 internal transcribed spacer region; (e) S2F and S3R to target the ITS2 internal transcribed spacer in terrestrial plants; (f) fITS7 or gITS7, and ITS4 to target the fungal ITS2 region; (g) NS31 and AML2 to target glomeromycota 18S rRNA genes; and (h) mICOIintF and jgHCO2198 to target cytochrome c oxidase subunit I (COI) genes in animals. More research is currently required to confirm primers suitable for the selective amplification of DNA from specific vertebrate taxa such as fish. Combined, these recommendations represent a framework for efficient, comprehensive and robust DNA-based investigations of biodiversity, applicable to most taxa and ecosystems. The adoption of standardised protocols for biodiversity assessment and monitoring using DNA extracted from environmental samples will enable more informative comparisons among datasets, generating significant benefits for ecological science and biosecurity applications

Changes in vegetation and soil characteristics in coastal sand dunes along a gradient of atmospheric nitrogen deposition

A field survey was conducted to detect signals of atmospheric nitrogen (N) in 11 dune systems along a nitrogen deposition gradient in the United Kingdom. In the mobile and semi-fixed dunes, above-ground biomass was positively related to N inputs. This increase was largely due to increased height and cover of Ammophila arenaria. In the long term, this increased biomass may lead to increased organic matter accumulation and consequently accelerated soil development. In the fixed dunes, above ground biomass also showed a positive relationship with N inputs as did soil C : N ratio while soil available N was negatively related to N inputs. Plant species richness was negatively related to N inputs. In the dune slacks, while soil and bulk vegetation parameters showed no relationship with N inputs, cover of Carex arenaria and Hypochaeris radicata increased. Site mean Ellenberg N numbers showed no relationship with N deposition either within habitats or across the whole dataset. Neither abundance-weighting nor inclusion of the Siebel numbers for bryophytes improved the relationship. The survey reveals that the relationships of soil and vegetation with atmospheric N deposition vary between sand dune habitats but, despite this variability, clear correlations with N inputs exist. While this survey cannot establish causality, on the basis of the relationships observed we suggest a critical load range of 10 - 20 kg N ha(-1) yr(-1) for coastal sand dunes in the UK

Abnormal Distracter Processing in Adults with Attention-Deficit-Hyperactivity Disorder

Background: Subjects with Attention-Deficit Hyperactivity Disorder (ADHD) are overdistractible by stimuli out of the intended focus of attention. This control deficit could be due to primarily reduced attentional capacities or, e. g., to overshooting orienting to unexpected events. Here, we aimed at identifying disease-related abnormalities of novelty processing and, therefore, studied event-related potentials (ERP) to respective stimuli in adult ADHD patients compared to healthy subjects. Methods: Fifteen unmedicated subjects with ADHD and fifteen matched controls engaged in a visual oddball task (OT) under simultaneous EEG recordings. A target stimulus, upon which a motor response was required, and non-target stimuli, which did not demand a specific reaction, were presented in random order. Target and most non-target stimuli were presented repeatedly, but some non-target stimuli occurred only once (‘novels’). These unique stimuli were either ‘relative novels ’ with which a meaning could be associated, or ‘complete novels’, if no association was available. Results: In frontal recordings, a positive component with a peak latency of some 400 ms became maximal after novels. In healthy subjects, this novelty-P3 (or ‘orienting response’) was of higher magnitude after complete than after relative novels, in contrast to the patients with an undifferentially high frontal responsivity. Instead, ADHD patients tended to smaller centro-parietal P3 responses after target signals and, on a behavioural level, responded slower than controls

Effectiveness of an online group course for adolescents and young adults with depressive symptoms: study protocol for a randomized controlled trial

<p>Abstract</p> <p>Background</p> <p>Depression is a common condition whose first onset is usually in late adolescence or early adulthood. Internet-based interventions are an effective treatment approach to depression. The aim of this study is to investigate the effectiveness of a Dutch online cognitive-behavioural group course known as Master Your Mood (<it>Grip op Je Dip</it>) for young people reporting depressive symptoms. Secondary research questions involve maintenance of effect at 6 months, mediators, and predictors of better outcomes.</p> <p>Methods</p> <p>We will conduct a randomised controlled trial (RCT) in which 244 young people aged 16-25 are randomly allocated to the Grip op Je Dip (GOJD) online group course or to a waiting list control group. The participants will be recruited from the general population. The primary outcome measure will be the severity of depressive symptoms according to the Center for Epidemiological Studies Depression Scale (CES-D). Other outcomes will include anxiety (Hospital Anxiety and Depression Scale-Anxiety, HADS) and mastery (Mastery Scale). Assessments will take place in both groups at baseline and three months later. Effect maintenance will be studied in the GOJD group six months after baseline, with missing data imputed using the expectation-maximisation method. Mediators and predictors of better outcomes will also be identified.</p> <p>Discussion</p> <p>The trial should add to the body of knowledge on the effectiveness of Internet-based interventions for depression. To our knowledge, this will be the first RCT on an online group intervention in this field.</p> <p>Trial registration</p> <p><a href="http://www.trialregister.nl/trialreg/admin/rctview.asp?TC=NTR1694">NTR1694</a></p